An important component of oncogenesis is the evasion of programmed cell death (also known as "apoptosis"), which is an essential mechanism of tumor suppression, as evasion of apoptosis is one of six acquired properties that result in a metastatic phenotype. Evasion of apoptosis offsets uncontrolled proliferation and can contribute to chemotherapeutic resistance, as many chemotherapeutic agents rely on induction of apoptosis in order to kill the cell. Apoptosis, and specifically the intrinsic mitochondrial pathway, relies on two Bcl-2 family proteins, Bax and Bak. Once activated, Bax and Bak oligomerize and produce mitochondrial outer membrane permeabilization, resulting in the release of intermembrane space proteins into the cytosol, thereby engaging downstream signaling cascades (such as caspase activation) that lead to the demise of the cell. To date, only three proteins are known to directly activate Bax and/or Bak: Bim, Bid and cytosolic p53. However, recent genetic and biochemical data suggest that mitochondrial apoptosis can proceed normally in the absence of these three proteins, indicating the existence of other "direct activators" of Bax and Bak. The goal of this proposal is to identify and characterize these non-canonical direct activators, and to explore their role in hematological cancers. Using genetic approaches, we will create cells that lack Bim, Bid and p53 expression in order to study the apoptotic response of cells lacking known direct activators of Bax and Bak. As a second approach to studying non-canonical direct activators, we will generate tumor cell lines from E?-myc mice lacking Bim and Bid expression, in order to examine whether such tumors contain functional non-canonical direct activators. Lastly, as a third approach, we will perform a large-scale screen for novel proteins that can act as direct activators. Loss of p53, Bim and Bid can contribute to carcinogenesis. It is therefore probable that loss of non-canonical direct activators can also contribute to cancer progression. Characterization of these proteins in hematological malignancies will enhance our knowledge of carcinogenesis, and provide researchers with important new therapeutic targets. Relevance: Cancer cells persist, in part, because they find ways to evade cell death, often through reducing expression of molecules that promote cell death, such as the proteins Bax and Bak and their activators, Bim, Bid and cytosolic p53. Recent genetic and biochemical evidence suggests proteins other than Bim, Bid and cytosolic p53 may exist, and if they do, they are likely dysregulated in cancer cells. Identifying and characterizing these non-canonical direct Bax/Bak activators and exploring their role in carcinogenesis will enhance our knowledge of carcinogenesis, and provide researchers with important new therapeutic targets.